以PCR及多位點酵素分析法探討自台灣地區分離之O157型大腸桿菌特性   全文下載

Characterization of Escherichia coli Serotype O157 Strains Isolated in Taiwan by PCR and Multilocus Enzyme Analysis

本研究自1997 年7 月至1999 年6 月，從台灣地區牛羊糞便或生肉檢體分離、蒐集O157 型大腸桿菌（Escherichia coli），共六株，僅有一株分離自羊糞檢體之菌株屬E. coli O157:H7 ，其餘為H41、H45、H?（以市售43 種O型血清無法鑑別出）、或不具運動性（non-motile, NM）。進一步以Polymerase chain reaction（PCR）方法分析屬Enterohemorrhagic E. coli（EHEC）之致病基因，僅該H7菌株具有slt2 、eaeA 及hlyA 等致病基因，同時該菌不能發酵山梨糖醇及缺乏β-D-葡萄糖啟酵素活性。五株non-H7 之O157 型菌株，再以PCR方法測試其他大腸桿菌致病基因，包括Enterotoxigenic E. coli（ETEC； heat-labile enterotoxin, LT； heatstable enterotoxin, ST）， Enteroinvasive E. coli（EIEC； invasive plasmid）及Enteropathogenic E. coli（EPEC；adherence factor, EAF），發現均不帶有前述之基因。為確定菌株之遺傳關係，選用十種酵素，進行多位點酵素分析（multilocus enzyme analysis, MLEA）。六株菌共區分成五種電泳圖譜型別（electrophoretic type, ET）。同時，以群集法分析菌株之純系關係，發現五株non-EHEC O157與EHEC O157:H7菌株明顯分屬於二個純系（clone）。根據結果推測，台灣本土分離之non-EHEC O157與EHEC O157:H7菌株可能衍生自二個純系，經由多位點酵素分析，可瞭解本土O157型大腸桿菌之遺傳關係。"

Escherichia coli serotype O157 strains were isolated from fecal samples of cattle and sheep, as well as raw meat in Taiwan from July 1997 to June 1999. In total, six strains of E. coli serotype O157 were collected. Of these, only one strain, isolated from fecal samples of sheep, was characterized as E. coli O157:H7. The remaining five strains were determined as H41, H45, H? (non-typeable with 43 O-antisera) and NM (non-motile). All six strains were analyzed for pathogenic genes of Enterohemorrhagic E. coli (EHEC) by the Polymerase chain reaction (PCR) method. The E. coli O157:H7 was the only strain harbored the virulent genes of slt2, eaeA and hlyA. Meanwhile, this strain was unable to ferment sorbitol and lacked β-D-glucuronidase activity. The five non-H7 serotype of E. coli O157 strains were further subjected to PCR analysis for other pathogenic genes, including genes in Enterotoxigenic E. coli (ETEC, heat-labile enterotoxin, LT, heatstable enterotoxin, ST), Enteroinvasive E. coli (EIEC, invasive plasmid) and Enteropathogenic E. coli (EPEC, adherence factor, EAF). Results revealed that the five non-H7 strains did not carry any of the above mentioned genes. To better understand the genetic relatedness among six isolated strains, ten enzyme loci were delineated using the multilocus enzyme analysis. Six strains were designated to five electrophoretic types. Cluster analysis further revealed that the five non-EHEC O157 and one EHEC E. coli O157:H7 strains belong to two distinct clones, respectively. These results suggested that non-EHEC O157 and EHEC E. coli O157:H7 strains isolated in the Taiwan area most likely originated from two clones. Furthermore, the genetic relatedness among indigenous E. coli O157 strains in Taiwan was elucidated through this multilocus enzyme analysis. 本研究自1997 年7 月至1999 年6 月，從台灣地區牛羊糞便或生肉檢體分離、蒐集O157 型大腸桿菌（Escherichia coli），共六株，僅有一株分離自羊糞檢體之菌株屬E. coli O157:H7 ，其餘為H41、H45、H?（以市售43 種O型血清無法鑑別出）、或不具運動性（non-motile, NM）。進一步以Polymerase chain reaction（PCR）方法分析屬Enterohemorrhagic E. coli（EHEC）之致病基因，僅該H7菌株具有slt2 、eaeA 及hlyA 等致病基因，同時該菌不能發酵山梨糖醇及缺乏β-D-葡萄糖啟酵素活性。五株non-H7 之O157 型菌株，再以PCR方法測試其他大腸桿菌致病基因，包括Enterotoxigenic E. coli（ETEC； heat-labile enterotoxin, LT； heatstable enterotoxin, ST）， Enteroinvasive E. coli（EIEC； invasive plasmid）及Enteropathogenic E. coli（EPEC；adherence factor, EAF），發現均不帶有前述之基因。為確定菌株之遺傳關係，選用十種酵素，進行多位點酵素分析（multilocus enzyme analysis, MLEA）。六株菌共區分成五種電泳圖譜型別（electrophoretic type, ET）。同時，以群集法分析菌株之純系關係，發現五株non-EHEC O157與EHEC O157:H7菌株明顯分屬於二個純系（clone）。根據結果推測，台灣本土分離之non-EHEC O157與EHEC O157:H7菌株可能衍生自二個純系，經由多位點酵素分析，可瞭解本土O157型大腸桿菌之遺傳關係。